The broad objective of the proposed research is to characterize the enzymatic reactivity of dissimilatory nitrite reductase. The specific goals are to determine a detailed steady state kinetic mechanism for the reactions of the enzyme, to determine the chemical events in the bimolecular reaction steps of the enzyme, and to determine the protein structural requirements for catalysis. Methods for the first goal include purification to homogeneity of the reductase from Paracoccus denitrificans and kinetic analysis of electron transport rates with donor proteins by spectrophotometric and polarographic techniques. For the second goal, rapid mixing solution kinetics will be followed for nitrite and other substrate analogs. For the third goal, reactivities will be analyzed in terms of active site geometries and a determination of the primary amino acid sequence in the active site domain will be made.